3772 Afr. J. Biotechnol.2004).
(African Curry plant) is a lowgrowing shrubby species in the family Labiatae. All thespecies in the genus contain strongly scented essentialoils. It is grown as pot herbs for local medicines andexists in diversity of forms and cultivars (Schippers,2000). The powders and essential oils of
species have been widely used for control of insect pestsespecially storage insect pests (Oparaeke et al
, 2002). Itis a slightly hairy annual with much branched angularstems carrying opposite, ovate leaves which are usuallyless than 1 cm long and borne on fairly long petioles. Ineastern Nigeria, it serves as a source of stimulant andcondiment in soup. Medicinally, it is used for thetreatment of stomach ache, sores and in management ofbabies cord after delivery among the Igbos of Nigeria(Ijeh et al
, 2004).The aim of the present study was to document thebiological effects of the essential oil on the maize weeviland to analyse its composition in order to assess itsrelative safety.
MATERIALS AND METHODS
was cultured in the laboratory at 27 ± 2ºC, 60 - 65%R.H and 12 h : 12 h light : dark regime. The food media used wasinsecticide-free whole maize grains purchased from a local market(Dikomba) outside Nairobi in Kenya. Fifty pairs of
wereplaced in 1 L glass jar containing 400 g of maize grains. The jarswere then covered with nylon mesh held in place with rubberbands. Grains were disinfested in the oven at 40
C for 4 h(Jembere et al
, 1995) and kept in the laboratory before use.
Plant collection and isolation of their essential oil
leaves were collected from Umudike, Nigeria inJanuary 2006. The identity of the plant was confirmed at theherbarium of Michael Okpara University of Agriculture, Umudike,Nigeria, before using them for the present study. Plants were air-dried in a well-ventilated area for five days before extraction.Voucher specimens are kept at the University herbarium.
The essential oil was extracted by steam distillation usingClavenger apparatus (Guenter, 1949). The condensing oils werecollected in n-hexane (Aldrich HPLC grade) and the solution wasfiltered and exposed to anhydrous sodium sulphate to remove anytraces of water. Hexane was then removed by distillation at 60
Cfrom 'Contes' Short Path distillation apparatus and the oil collectedand weighed, and stored in small amber-coloured vials.
Analysis of essential oil
Gas chromatographic (GC) analyses were performed on a capillarygas-chromatograph Hewlett Packard (HP) 5890 Series II equippedwith a split-less capillary injector system, 50 m x 0.20 mm (i.d.)cross-linked with HP-ultra 1methylsilicone 0.33 µm (film thickness)capillary column, and Flame Ionization Detector (FID) coupled toHP 3393A Series II integrator. The integrator was used to calculatethe peak areas. The carrier gas was nitrogen at a flow rate of 0.84ml/min. The temperature programme comprised of an initialtemperature of 40
C (0 min) to 90
C at 7
C/min, a hold at thistemperature for 5 min, then to 115 at 3
C/min followed by anotherhold for 5 min, and finally to 280
C at 4
C/min where it wasmaintained for 20 min.Gas chromatography–linked mass Spectrometry (GC-MS)analysis was carried out on a HP 8060 Series II gas chromatographcoupled to VG Platform II Mass Spectrometer (manufactured byMicromass, UK, formerly known as VG Biotech). The MS wasoperated in the Electron impact mode (EI) at 70 eV and anemission current of 200
A. The temperature of source was held at180
C and the multiplier voltage at 300 V. The pressure of the ionsource and MS detector were held at 9.4 x 10
and 9.4 x 10
mbar,respectively. The MS had a scan cycle of 1.5 s (scan duration of 1 sand inter-scan delay, 0.5 s). The mass and scan range was set atm/z 1 - 1400 and 38 - 650, respectively. The instrument wascalibrated using heptacosafluorotributyl amine, [CF
N,(Apollo Scientific Ltd., UK). Column (film thickness 0.5
m)temperature was programmed as in the case of GC analysis. AllGC-MS analyses were made in the splitless mode with helium asthe carrier gas. Preliminary identification of constituents was basedon computer matching components of mass spectral data againstthe standard Wiley and NIST library spectra, constituted fromspectra of pure substances and components of the known essentialoils, and literature MS data. They were confirmed by their GCretention time comparison with those of reference compounds,peak enhancement as well as co-injection /co-elution with authenticsamples. The samples used were obtained from Aldrich ChemicalsUK. Relative proportion of the essential oil was computed in eachcase from GC-MS peak areas.
Repellent action of the essential oil against
wasassessed in a choice bioassay system at 27 ± 2
C and 60 - 65%R.H. previously reported by Bekele et al. (1996). It consisted oftwo 1 L glass jars connected together at their rims by means of a30 x 10 cm nylon mesh tube. A 5.0 cm diameter circular hole wascut at the middle of the mesh for the introduction of test insects.250 g of maize were put into each glass jars. Grains in one jarwere treated with the essential oil at the rate of 0.012, 0.06 and0.3% (30, 150 and 750 mg/250 g) while untreated grains in theother jar acted as control. Twenty-five adults of
introduced into the nylon mesh tube through the circular hole bymeans of a 5 cm diameter funnel. The number of insects presentat the control (N
) and treated (N
) jars were recorded after 1-hour exposure. All repellency assays were replicated four times.Percent repellency (PR) values were computed from the formula:PR = [N
] × 100.PR data were analyzed using ANOVA after arcsinetransformation.
Effect of essential oil on mortality
Essential oil was applied to the grains at the rate of 0.012, 0.06and 0.3% dissolved in 10 ml of 95% n-hexane and shakenthoroughly to ensure uniform distribution over grain surface.Treated grains were kept for 24 h to allow the hexane to evapo-rate completely before bioassays were conducted. Two blankcontrols were run concurrently consisting of hexane treated grainand untreated grain. Ten pairs of 5/7-day-old